CN105368800B - A kind of thermal starting Taq DNA polymerase and preparation method thereof - Google Patents

A kind of thermal starting Taq DNA polymerase and preparation method thereof Download PDF

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CN105368800B
CN105368800B CN201510891376.0A CN201510891376A CN105368800B CN 105368800 B CN105368800 B CN 105368800B CN 201510891376 A CN201510891376 A CN 201510891376A CN 105368800 B CN105368800 B CN 105368800B
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刘未斌
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Abstract

The present invention provides a kind of thermal starting Taq archaeal dna polymerase and preparation method thereof.The thermal starting Taq archaeal dna polymerase is obtained in conjunction with aldehyde radical polysaccharide by Taq archaeal dna polymerase; the combination reacts to form amido bond realization with the aldehyde radical of aldehyde radical polysaccharide by the amino of Taq archaeal dna polymerase; it disconnects within the amide bond energy 5 minutes or more at a temperature of 95 DEG C; to discharge the polymerization activity of Taq archaeal dna polymerase; polysaccharide has protective effect to Taq archaeal dna polymerase simultaneously; not only inhibited Taq archaeal dna polymerase at low temperature 3 ' to 5 ' polymerization activity, but also protective agent as Taq archaeal dna polymerase is conducive to enhance the efficiency of PCR reaction.Molecular biology, the fields such as forensic dna detection can be widely used using thermal starting Taq archaeal dna polymerase prepared by this method.

Description

A kind of thermal starting Taq DNA polymerase and preparation method thereof
Technical field
The present invention relates to archaeal dna polymerase and preparation method thereof more particularly to a kind of thermal starting Taq archaeal dna polymerase and its systems Preparation Method.
Background technique
PCR reaction (Polymerase Chain Reaction) is the beyond body nucleic acid amplification that the mid-80 grows up Technology.Its purpose is that have special, sensitive, yield height so that DNA segment expands rapidly in a short time, quickly, easy, It is reproducible, it is easy to outstanding advantages of automating, it is made to have obtained extensive reality rapidly in molecular biology and medical domain Using.It is considered as the most important technological break-through of molecular biology field in recent decades by many scientists.
Round pcr has started a revolution in life science, it can make people invisible spectro by several hours DNA polymerization reaction, so that it may by DNA cloning 109Times.The DNA being present in sample can be not only expanded by round pcr, it can also To be enriched with any one of hybrid dna molecule.The important value of PCR is amplification, and there are micro and special DNA sequence dnas.This Technology is to be invented by Kary Mullis in 1985, he obtains Nobel chemistry Prize in 1993 thus.
The principle of PCR:
The semi-conservative replication of DNA is the important channel of biological evolution and passage.Double-stranded DNA can be under the action of a variety of enzymes Unwinding is denaturalized at single-stranded, in the presence of archaeal dna polymerase and promoter, is copied into same two according to base pair complementarity principle Molecule copy.Find in an experiment, DNA can also occur at high temperature be denaturalized unwinding, when temperature reduce after again can with renaturation at For double-strand.Therefore, the denaturation and renaturation of DNA are controlled by temperature change, and design primer does promoter, add human DNA polymerase, DNTP can complete the replication in vitro of specific gene.
The PCR process of standard is divided into three steps:
1.DNA is denaturalized (90 DEG C -96 DEG C): for double-stranded DNA template under heat effect, hydrogen bond fracture forms single stranded DNA.
2. annealing (8 DEG C -65 DEG C): system temperature reduces, and primer forms local double-strand in conjunction with DNA profiling.
3. extending (70 DEG C -75 DEG C): under the action of Taq DNA polymerase (in 72 DEG C or so optimal activity), with DNTP is raw material, holds from the 3 ' of primer to 5 ' ends and extends, synthesizes the DNA chain complementary with template.
In round pcr, heat-resisting TAQ archaeal dna polymerase is a key.The application value of Taq DNA polymerase according to Rely the functional characteristic in it, main functional characteristics have:
(1) heat resistance: hot resistant DNA polymerase is the key that polymerase chain reaction (PCR) technology is achieved automation. It makes PCR process high temperature (90 DEG C or more) unwinding, (60 DEG C or so) of low temperature annealing and (about 70 DEG C) of medium temperature extensions can be in enzyme Iterative cycles in the case where being added at one time.
(2) polymerization activity and efficiency: refer to enzyme molecule to the compatibility of template and substrate and the catalytic efficiency of enzyme, generally It is indicated with converting number and highest than work.Template and concentration of substrate, buffer system and divalent ion and amplification target in reaction system The length of DNA also has large effect to polymerization activity and catalytic efficiency.
(3) 3 ' -5 ' 5 prime excision enzyme activities: the archaeal dna polymerase having is exo-acting with 3 ' -5 ', has TAQ archaeal dna polymerase There are 3 ' end mistakes in proofreading or corrigendum duplication or repair process.
(4) 5 ' -3 ' 5 prime excision enzyme activities: the archaeal dna polymerase having is exo-acting with 5 ' -3 ', it can be used in PCR product and examine Examining system, to generate the signal of the detectable amplification of specificity.But the hydrolysis of the plasmid template to template especially cyclic DNA Effect will affect the amplification efficiency of PCR.
(5) reverse transcriptase activity: the archaeal dna polymerase having has reverse transcriptase activity, and especially heat-resisting has reverse transcriptase activity Archaeal dna polymerase, can be used for cDNA library single enzyme process building and amplification, mRNA secondary structure can be overcome to transcriptive process,reversed Inhibiting effect simplifies building and amplification procedure.
(6) fidelity: mainly related with compatibility and recognition capability of the enzyme to template and substrate.Followed by 3 ' -5 ' outside The corrective action cut is greatly improved the fidelity of archaeal dna polymerase.High fidelity is conducive to expand long segment DNA, fidelity compared with It is easy to stop to expand in fault out when low.
Taq enzyme is a kind of heat resistance list subunit enzyme, and molecular weight 93909Da is made of 832 amino acid, containing there is 7 Structural domain, functional area are positioned between 287-832 amino acid.Zymetology is classified as EC1.7.7.7, has 5 ' -3 ' end polymerizations Enzyme characteristic, optimum temperature are 72-80 DEG C, and 95 DEG C of half-life period are 40min, and 100 DEG C of half-life period are 5min.Higher than DNA denaturation temperature, It can be consequently used for PCR amplification.Taq archaeal dna polymerase is highest one kind of activity in the hot resistant DNA polymerase found so far, often Molecule enzyme each second may extend away 150 liquor-saturated acid of core, be that extension speed is most fast, the DNA of amplification in all thermophilic archaeal dna polymerases Length is up to 20kb.
Thermal starting polymerase chain reaction (Hot start PCR) is a kind of polymerase chain reaction method of improvement, main To be used to the amplification for avoiding generating non-specific sequences in operating process.Archaeal dna polymerase and other reactants are completely cut off, or made The polymerase that can just activate under high fever situation avoids beginning to react before not up to set temperature.PCR heat is realized at present Starting method includes: that manual hot start method, the pack of wax protective layer, antibody modification method and chemical modification method etc. are several.Quotient Industry thermal starting Taq archaeal dna polymerase further includes based on antibody modification method and chemical modification method etc..Manual thermal starting is cumbersome, Infeasible in practical applications, wax protective layer pack is highly unstable to be commercialized, and the antibody that antibody act uses is to pass through The antibody that animal immune obtains, preparation process is complicated, expensive.
How the thermal starting Taq DNA polymerization that one kind is simple, effective preparation has stable enhancing PCR reaction efficiency is provided The method of enzyme becomes problem to be solved.
Summary of the invention
The present invention provides a kind of thermal starting Taq archaeal dna polymerase, and the Taq archaeal dna polymerase is by Taq DNA polymerase and aldehyde Base polysaccharide, which combines, to be obtained, and the combination reacts shape with the aldehyde radical of aldehyde radical polysaccharide by the amino of Taq archaeal dna polymerase It realizes, disconnects at amido bond within the amide bond energy 5 minutes or more at a temperature of 95 DEG C, to discharge the poly- of Taq archaeal dna polymerase Close activity, while polysaccharide has protective effect to albumen, both inhibited Taq archaeal dna polymerase at low temperature 3 ' to 5 ' polymerization it is living Property, and the protective agent as Taq archaeal dna polymerase is conducive to enhance the efficiency of PCR reaction.
The present invention also provides a kind of method for preparing thermal starting Taq archaeal dna polymerase, by the method can it is simple, have The preparation of effect has the thermal starting Taq archaeal dna polymerase for stablizing enhancing PCR reaction efficiency.
A kind of thermal starting Taq archaeal dna polymerase provided by the invention, the Taq archaeal dna polymerase is by Taq archaeal dna polymerase It is obtained in conjunction with aldehyde radical polysaccharide, the combination is anti-by the aldehyde radical of the amino and aldehyde radical polysaccharide of Taq archaeal dna polymerase Amido bond realization should be formed, is disconnected within the amide bond energy 5 minutes or more at a temperature of 95 DEG C, the polysaccharide is expressed from the next, Wherein n is 40-200,
In the scheme of the application, the Taq archaeal dna polymerase for being used to prepare the application thermal starting Taq archaeal dna polymerase can be with Selected from commonly used in the art, commercially available general T aq archaeal dna polymerase, quick Taq archaeal dna polymerase, high-fidelity Taq archaeal dna polymerase and long segment amplification Taq archaeal dna polymerase etc..Further, enzyme activity unit can be greater than 150U/mg.
In the specific embodiment of the present invention, the aldehyde radical polysaccharide is obtained by oxidizing polysaccharide, The oxidant is NaIO4。
In the scheme of the application, the polysaccharide is glucan or chitosan.Further, the polysaccharide is poly- for Portugal Sugar 40, macrodex or chitosan 50.
Further, the condition of the oxidizing polysaccharide are as follows: polysaccharide described in 1g-10g is dissolved in 100mL's The NaIO of 0.02M-0.2M4Acetate buffer in, the pH of the buffer is 4-6, is then protected from light 20h- at 4 DEG C -8 DEG C 48h is dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powdered aldehyde radical polysaccharide.
Further, the condition of the oxidizing polysaccharide are as follows: polysaccharide described in 2-5g is dissolved in 100mL's The NaIO of 0.1M-0.15M4Acetate buffer in, the pH of the buffer is 4-6, is then protected from light 20h- at 4 DEG C -8 DEG C 48h is dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powdered aldehyde radical polysaccharide.
Further, condition of the Taq archaeal dna polymerase in conjunction with aldehyde radical polysaccharide are as follows: the aldehyde radical is more Glycan is added in the aqueous solution for the Taq archaeal dna polymerase that concentration is 50mg/L and forms oxidation mixture, wherein the aldehyde radical is more The molar ratio of glycan and Taq archaeal dna polymerase is 100-150:1, and 4 DEG C -8 DEG C are protected from light 20h-48h, based on mixing after reaction The gross weight of object is added the bovine serum albumin (BSA) that 1-5wt% concentration is 10wt%, closing is then protected from light at 4 DEG C -8 DEG C 20h-48h separates the acquisition thermal starting Taq archaeal dna polymerase of generation.The progress of this field conventional means can be used in the separation, Such as dialyse, post separation or ultrafiltration etc..Further, in the solution of the present invention, the aldehyde radical polysaccharide and Taq DNA are poly- The molar ratio of synthase is 100-130:1.
A kind of method preparing thermal starting Taq archaeal dna polymerase provided by the invention, by Taq archaeal dna polymerase by Taq Archaeal dna polymerase obtains thermal starting Taq archaeal dna polymerase in conjunction with aldehyde radical polysaccharide, and the combination passes through Taq archaeal dna polymerase Amino reacted with the aldehyde radical of aldehyde radical polysaccharide to be formed amido bond realization, the amide bond energy at a temperature of 95 DEG C 5 minutes with Upper disconnection, the polysaccharide are expressed from the next, and wherein n is 40-200,
In the specific embodiment of the present invention, the aldehyde radical polysaccharide is obtained by the oxidizing polysaccharide , the oxidant is NaIO4。
Further, the polysaccharide is glucan or chitosan.
Further, the condition of the oxidizing polysaccharide are as follows: polysaccharide described in 1g-10g is dissolved in 100mL's The NaIO of 0.02M-0.2M4Acetate buffer in, the pH of the buffer is 4-6, is then protected from light 20h- at 4 DEG C -8 DEG C 48h is dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powdered aldehyde radical polysaccharide.
Further, the condition of the oxidizing polysaccharide are as follows: polysaccharide described in 2-5g is dissolved in 100mL's The NaIO of 0.1M-0.15M4Acetate buffer in, the pH of the buffer is 4-6, is then protected from light 20h- at 4 DEG C -8 DEG C 48h is dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powdered aldehyde radical polysaccharide.
Further, condition of the Taq archaeal dna polymerase in conjunction with aldehyde radical polysaccharide are as follows: the aldehyde radical is more Glycan is added in the aqueous solution that concentration is 50mg/L Taq archaeal dna polymerase and forms oxidation mixture, wherein the aldehyde radical poly The molar ratio of sugar and Taq archaeal dna polymerase is 100-150:1, and 4 DEG C -8 DEG C are protected from light 20h-48h, then mix after being based on reaction The gross weight of object is added the bovine serum albumin that 1-5wt% concentration is 10wt% and is protected from light closing 20h-48h at 4 DEG C -8 DEG C, point Thermal starting Taq archaeal dna polymerase from generation.
The present invention program has the advantage that
1) the amide bond energy in thermal starting Taq archaeal dna polymerase of the invention breaks for 5 minutes or more under 95 DEG C of high temperature actions It opens, so that the polymerization activity of Taq archaeal dna polymerase is discharged, due to its no activity before the thermal denaturation of initial cycle, thus Inhibit under cryogenic conditions as primer non-specificity anneal or primer dimer caused by non-specific amplification, have when amplification general The activity of logical high temperature-resisting DNA polymerase, while there is stronger specificity.
2) thermal starting Taq archaeal dna polymerase of the invention can be widely applied to high sensitivity and have stronger background genes group to expand Increase (detections of some specific gene site or foreign pathogens in such as genome), quantitative fluorescent PCR, determined dna sequence, Multiplex PCR, TA clone etc..
3) preparation thermal starting Taq archaeal dna polymerase method provided by the invention simply, can be prepared effectively to have to stablize and be increased The thermal starting Taq archaeal dna polymerase of strong PCR reaction efficiency.
Detailed description of the invention
Fig. 1 is aldehyde radical polysaccharide molecular formula；
Fig. 2 is the structural schematic diagram of thermal starting Taq archaeal dna polymerase of the present invention；
Fig. 3 A is the agarose electrophoresis figure obtained using non-thermal starting enzymatic amplification human genome；Fig. 3 B is using the present invention Thermal starting Taq archaeal dna polymerase amplification human genome obtain agarose electrophoresis figure；
It is limited that Fig. 4 is that the thermal starting Taq archaeal dna polymerase prepared using the method for the present invention expands the general safe science and technology of Beijing Alioth The parting map of the amplified production obtained after company's Inteligene Search C kit.
Specific embodiment
Embodiment 1
The preparation of thermal starting Taq archaeal dna polymerase of the present invention
1, it prepares aldehyde radical polysaccharide: Gentran 40 (molecular weight 40kDa) 2.5g is dissolved in NaIO4The vinegar of (0.12M) In phthalate buffer (pH 6) 100ml, it is protected from light in 4 DEG C for 24 hours, is dialysed 3 times with distilled water, it is concentrated, it is powderedly dry Aldehyde radical glucan；Fig. 1 is aldehyde radical polysaccharide molecular formula.
2, the combination of Taq archaeal dna polymerase and aldehyde radical polysaccharide:
By powdered aldehyde radical Dextran 8 mg, the 50mg/L Taq archaeal dna polymerase of the Promga company production of 3ml is added Oxidation mixture is formed in the aqueous solution of (molecular weight 75kDa), enzyme activity unit is greater than 150U/mg, and 4 DEG C are protected from light for 24 hours, then Based on the gross weight of mixture after reaction, the bovine serum albumin BSA that the mass fraction for being added 1% is 10wt% is protected from light anti-at 4 DEG C It should close for 24 hours；
The reaction mixture for the archaeal dna polymerase containing Taq that reaction is completed, upper 1.6 × 55cm, Sephadex G100 column Son is operated according to Sephadex G100 column operation instructions, and adjusting flow velocity is 0.3ml/min, is collected first and is greater than The destination protein peak of 90KD is added and saves buffer (50mmol Tris-HCl (PH, 8.0), 50mmol KCl, 0.1mmol EDTA, 1mmolDTT, 0.5mmol PMSF, 50% glycerol) to get thermal starting Taq archaeal dna polymerase.Fig. 2 is the heat of the application Start Taq DNA polymerase structural schematic diagram.
Embodiment 2
The preparation of thermal starting Taq archaeal dna polymerase of the present invention
1, prepare aldehyde radical polysaccharide: macrodex (molecular weight 70kDa) 4.375g is dissolved in NaIO4The vinegar of (0.12M) In phthalate buffer (pH4) 100ml, it is protected from light in 4 DEG C for 24 hours, is dialysed 4 times with distilled water, concentration, dry powdered aldehyde radical Change glucan；
2, the combination of Taq archaeal dna polymerase and aldehyde radical polysaccharide:
By powdered aldehyde radical glucan 14mg, the 50mg/L high-fidelity Taq DNA of the Takara company production of 3ml is added Oxidation mixture is formed in the aqueous solution of polymerase (molecular weight 90kDa), enzyme activity unit is greater than 150U/mg, and 4 DEG C are protected from light For 24 hours, then the gross weight based on mixture after reaction, the BSA that the mass fraction for being added 1% is 10wt% are protected from light envelope at 8 DEG C Close 30h；
Will reaction complete the archaeal dna polymerase containing Taq reaction mixture, with molecular cut off be 70KD super filter tube into Row is concentrated, 8000rpm centrifugation 30min under the conditions of 4 DEG C, concentrate preservation buffer (50mmolTris-HCl (PH, 8.0), 50mmol KCl, 0.1mmol EDTA, 1mmolDTT, 0.5mmol PMSF) buffer restores to reaction mixture original volume, adds Enter with isometric glycerol, mix to get thermal starting Taq archaeal dna polymerase.Fig. 2 is that the thermal starting Taq DNA of the application polymerize Enzymatic structure schematic diagram.
Embodiment 3
The preparation of thermal starting Taq archaeal dna polymerase of the present invention
1, prepare aldehyde radical polysaccharide: chitosan 50 (molecular weight 50kDa) 3.125g is dissolved in NaIO4The vinegar of (0.12M) In phthalate buffer (pH4-6) 100ml, it is protected from light in 4 DEG C for 24 hours, is dialysed 3 times with distilled water, concentration, dry powdered aldehyde Base chitosan；
2, the combination of Taq archaeal dna polymerase and aldehyde radical polysaccharide:
By powdered aldehyde radical chitosan 10mg, the quick Taq DNA of 50mg/L of the Promega company production of 3ml is added Oxidation mixture is formed in the aqueous solution of polymerase (molecular weight 94kDa), enzyme activity unit is greater than 150U/mg, and 4 DEG C are protected from light For 24 hours, then the gross weight based on mixture after reaction, the BSA that the mass fraction for being added 1% is 10wt% are protected from light envelope at 6 DEG C Close 40h；
The reaction mixture for the archaeal dna polymerase containing Taq that reaction is completed is 70KD dialysis membrane with molecular cut off, 4 DEG C With preservation buffer (50mmol Tris-HCl (PH, 8.0), 50mmol KCl, 0.1mmol EDTA, 1mmolDTT, 0.5mmol PMSF), dialyse 48h, and addition and isometric glycerol mix to get thermal starting Taq archaeal dna polymerase.Fig. 2 is the heat of the application Start Taq archaeal dna polymerase structural schematic diagram.
Embodiment 4
Respectively using the thermal starting Taq archaeal dna polymerase of the application and non-thermal starting Taq archaeal dna polymerase, the base of people is expanded Because of a group DNA (source: 9947A promega company), comprising the following steps:
1, the genomic DNA of human blood is extracted using omega poba gene group extracts kit；
2, said gene group DNA is expanded on ABI9700PCR instrument, amplification system are as follows:
Amplification condition are as follows:
3, electrophoretic analysis is carried out using 1.5% agarose to the DNA amplification that amplification obtains, as a result as shown in Figure 3.
Fig. 3 A shows the agarose electrophoresis figure obtained using non-thermal starting enzymatic amplification human genome；Fig. 3 B, which is shown, to be made The agarose electrophoresis figure obtained with the thermal starting Taq archaeal dna polymerase amplification human genome of the application, it can be seen that You Benfa Bright method preparation thermal starting Taq archaeal dna polymerase is compared to non-thermal starting Taq archaeal dna polymerase, it is suppressed that by drawing under cryogenic conditions Non-specific amplification caused by the annealing of object non-specificity or primer dimer, with common high temperature-resisting DNA polymerase when amplification Activity, while there is stronger specificity.
Embodiment 5
Beijing Alioth Pu Tai Science and Technology Ltd. Inteligene is expanded using the thermal starting Taq archaeal dna polymerase of the application Search C kit, is carried out according to kit specification amplification step, and the product after amplification is analyzed through genetic analyzer and obtained Parting profiling results as shown in figure 4, explanation by the method for the present invention preparation thermal starting Taq archaeal dna polymerase can be good at expanding Multiple fluorescence PCR obtains corresponding target fragment.

Claims (8)

1. a kind of thermal starting Taq archaeal dna polymerase, the thermal starting Taq archaeal dna polymerase is by Taq archaeal dna polymerase and aldehyde radical Polysaccharide, which combines, to be obtained, and the combination reacts to form acyl by the amino of Taq archaeal dna polymerase with the aldehyde radical of aldehyde radical polysaccharide Amine key is realized, is disconnected within the amide bond energy 5 minutes or more at a temperature of 95 DEG C, and the polysaccharide is Gentran 40, macrodex Or chitosan 50.
2. thermal starting Taq archaeal dna polymerase according to claim 1, the aldehyde radical polysaccharide is by oxidizing described Polysaccharide obtains, and the oxidant is NaIO4。
3. thermal starting Taq archaeal dna polymerase according to claim 2, the condition of the oxidizing polysaccharide are as follows: will Polysaccharide described in 1g-10g is dissolved in the NaIO of the 0.02M-0.2M of 100mL4Acetate buffer in, the pH of the buffer is Then 4-6 is protected from light 20h-48h at 4 DEG C -8 DEG C, dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powder The aldehyde radical polysaccharide of last shape.
4. thermal starting Taq archaeal dna polymerase according to claim 1, the Taq archaeal dna polymerase and aldehyde radical polysaccharide In conjunction with condition are as follows: by the aldehyde radical polysaccharide be added concentration be 50mg/L Taq archaeal dna polymerase aqueous solution in formed Oxidation mixture, wherein the molar ratio of the aldehyde radical polysaccharide and Taq archaeal dna polymerase is 100-150:1,4 DEG C -8 DEG C are protected from light 20h-48h, then the gross weight based on mixture after reaction are reacted, the bovine serum albumin that 1-5wt% concentration is 10wt% is added, then It is protected from light closing 20h-48h at 4 DEG C -8 DEG C, separates the thermal starting Taq archaeal dna polymerase of generation.
5. a kind of method for preparing thermal starting Taq archaeal dna polymerase obtains Taq archaeal dna polymerase in conjunction with aldehyde radical polysaccharide Thermal starting Taq archaeal dna polymerase, the combination react shape with the aldehyde radical of aldehyde radical polysaccharide by the amino of Taq archaeal dna polymerase It realizes, disconnects at amido bond within the amide bond energy 5 minutes or more at a temperature of 95 DEG C, the polysaccharide is Gentran 40, Portugal is poly- Sugar 70 or chitosan 50.
6. described according to the method described in claim 5, the aldehyde radical polysaccharide is obtained by the oxidizing polysaccharide Oxidant is NaIO4。
7. according to the method described in claim 6, the condition of the oxidizing polysaccharide are as follows: by polysaccharide described in 1g-10g It is dissolved in the NaIO of the 0.02M-0.2M of 100mL4Acetate buffer in, the pH of the buffer is 4-6, then at 4 DEG C -8 DEG C it is protected from light 20h-48h, is dialysed 3-4 times with distilled water after reaction, concentrated to be dried to obtain powdered aldehyde radical more Glycan.
8. according to the method described in claim 5, condition of the Taq archaeal dna polymerase in conjunction with aldehyde radical polysaccharide are as follows: will The aldehyde radical polysaccharide is added in the aqueous solution for the Taq archaeal dna polymerase that concentration is 50mg/L and forms oxidation mixture, wherein The molar ratio of the aldehyde radical polysaccharide and Taq archaeal dna polymerase is 100-150:1, and 4 DEG C -8 DEG C are protected from light 20h-48h, then Based on the gross weight of mixture after reaction, the bovine serum albumin that 1-5wt% concentration is 10wt% is added and is protected from light at 4 DEG C -8 DEG C 20h-48h is closed, the thermal starting Taq archaeal dna polymerase of generation is separated.
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